IL22 Inhibits Epithelial Stem Cell Expansion in an Ileal Organoid Model

Abstract

Background & AimsCrohn’s disease is an inflammatory bowel disease that affects the ileum and is associated with increased cytokines. Although interleukin (IL)6, IL17, IL21, and IL22 are increased in Crohn’s disease and are associated with disrupted epithelial regeneration, little is known about their effects on the intestinal stem cells (ISCs) that mediate tissue repair. We hypothesized that ILs may target ISCs and reduce ISC-driven epithelial renewal.MethodsA screen of IL6, IL17, IL21, or IL22 was performed on ileal mouse organoids. Computational modeling was used to predict microenvironment cytokine concentrations. Organoid size, survival, proliferation, and differentiation were characterized by morphometrics, quantitative reverse-transcription polymerase chain reaction, and immunostaining on whole organoids or isolated ISCs. ISC function was assayed using serial passaging to single cells followed by organoid quantification. Single-cell RNA sequencing was used to assess Il22ra1 expression patterns in ISCs and transit-amplifying (TA) progenitors. An IL22-transgenic mouse was used to confirm the impact of increased IL22 on proliferative cells in vivo.ResultsHigh IL22 levels caused decreased ileal organoid survival, however, resistant organoids grew larger and showed increased proliferation over controls. Il22ra1 was expressed on only a subset of ISCs and TA progenitors. IL22-treated ISCs did not show appreciable differentiation defects, but ISC biomarker expression and self-renewal–associated pathway activity was reduced and accompanied by an inhibition of ISC expansion. In vivo, chronically increased IL22 levels, similar to predicted microenvironment levels, showed increases in proliferative cells in the TA zone with no increase in ISCs.ConclusionsIncreased IL22 limits ISC expansion in favor of increased TA progenitor cell expansion.

The role of IL-22 in intestinal health and disease

“…ISCs undergo asymmetric division to self-renew while generating committed transit-amplifying cells, which in turn further differentiate to give rise to all cell types found in the intestinal epithelium (Gehart and Clevers, 2019). IL-22RA1 is abundantly expressed across intestinal epithelial lineages (Lindemans et al, 2015; Zwarycz et al, 2018), and stimulation with IL-22 increases intestinal organoid proliferation and expands organoid size in vitro. IL-22 transgenic mice have an increased number of Ki67 + IECs and intestinal crypt cells and show increased crypt height (Zwarycz et al, 2018).…”

“…IL-22RA1 is abundantly expressed across intestinal epithelial lineages (Lindemans et al, 2015; Zwarycz et al, 2018), and stimulation with IL-22 increases intestinal organoid proliferation and expands organoid size in vitro. IL-22 transgenic mice have an increased number of Ki67 + IECs and intestinal crypt cells and show increased crypt height (Zwarycz et al, 2018). Mechanistically, IL-22 and STAT3 signaling promote epithelial cell survival upon radiation or epithelial damage induced by methotrexate (Aparicio-Domingo et al, 2015; Lindemans et al, 2015; Pickert et al, 2009), as STAT3 directly induces prosurvival genes, including B cell lymphoma ( BCL xl ), MCL1 , and Hsp70 (Grivennikov et al, 2009; Pickert et al, 2009).…”

“…It should be noted that the precise target cell type of IL-22 in the intestinal epithelium is a matter of ongoing debate. Two recent papers suggested that the mechanism by which IL-22 promotes regeneration in vivo and organoid growth in vitro involves stimulation of transit-amplifying cells, while it actually suppresses ISC expansion through activation of Wnt and Notch pathways (Zha et al, 2019; Zwarycz et al, 2018). Additional work is required to reconcile these seemingly contradictory findings.…”

IL-22 receptor signaling in Paneth cells is critical for their maturation, microbiota colonization, Th17-related immune responses, and anti-Salmonella immunity

“…14,26 Likewise, IL-22 is a key player in mucosal host defense. [1][2][3][4][5][6] All intestinal epithelial lineages including Paneth cells and ISCs express the IL-22 receptor; 19 however, it is not known whether IL-22-dependent regulation of small intestinal host defense is dependent on ISCs or a specific epithelial lineage. To examine differences in gene expression among small intestinal enterocytes and secretory cells, single-cell RNA sequencing (scRNA-seq) was performed on naive C57BL/6 mouse-derived primary small intestinal organoids.…”

<p>Curcumin Modifies Epithelial–Mesenchymal Transition in Colorectal Cancer Through Regulation of miR-200c/EPM5</p>

“…In line with the previous studies, a luciferase reporter of IL6 was also downregulated by miR-200c. 30,31 Taken together, miR-200c directly repressed EPM5 through the binding site in its 3ʹ UTR.…”